Dissertations / Theses on the topic 'Meningitis'

Infecções por Neisseria meningitidis estão associadas a altos índices de morbimortalidade no mundo. Na Região da Grande São Paulo a Doença Meningocócica (DM) causada por Neisseria meningitidis sorogrupo C começou a se tornar prevalente em 2001 , representando, em 2003, 62,7% de todos os casos de DM sorogrupados sendo que aproximadamente 88,5% dessas cepas eram não sorotipados e não sorosubtipados (C:NST:NSST). Estes dados sugeriam que um fenótipo, C: NST:NSST, tinha emergido na Grande São Paulo e, considerando-se a importância histórica da doença na região, iniciamos o presente estudo com o objetivo de esclarecer a mudança na dinâmica da DM pela determinação das características fenotípicas e genotípicas destas cepas. Para tanto, analisamos por sorotipagem, tipagem das regiões Variáveis da PorA e PorB e do gene 16S RNA ribossomal, 753 cepas de N. meningitidis C isoladas de casos de DM provenientes da Grande São Paulo, no período de 1990 a 2003. Dado a impossibilidade de caracterização do novo fenótipo pelos anticorpos monoclonais disponíveis mundialmente, objetivamos também a produção de hibridomas produtores desses anticorpos para caracterização do fenótipo C:NST:NSST. Foram selecionadas duas linhagens celulares híbridas, produtoras de anticorpos monoclonais que reconhecem as proteínas PorA e PorB deste novo fenótipo. Entre as 255 cepas de N. meningitidis C inicialmente caracterizadas como NST:NSST, 75% (n=191) tomaram-se completamente sorotipadas como 23:P1.14-6. A análise da similaridade do gene 16S RNA ribossomal das cepas analisadas demonstrou um único padrão genético, sugerindo a clonalidade deste novo fenótipo. Os dados obtidos neste trabalho, demonstram a introdução, na Região da Grande São Paulo, de um novo clone de Neisseria meningitidis C apresentando o fenótipo C:23:P1.14-6 e que está sendo responsável pelo aumento dos casos de DM causada por este sorogrupo.
Neisseria meningitidis (Men) is an important cause of morbidity and mortality and is a leading cause of bacterial meningitis and septicemia in children and young adults in Brazil. Meningococcal disease caused by MenC started becoming the most prevalent serogroup in 2001, representing 62.7% of all MD cases serogrouped in 2003 in Greater Sao Paulo and approximately 88.5% of MenC isolates were nonserotypeable and non-serosubtypeable (NST:NSST). This data suggested that a novel invasive isolate (C:NT:NSST) had emerged in GSP, and considering the historical importance of MenC disease in the region, we initiated this study to better understand the dynamics of MD looking at the phenotypic and molecular characteristics of these isolates. To accomplish this goal, we characterized 753 MenC isolates recovered during the period of 1990 to 2003 by serotyping, PorS and PorA VR typing, 16S rRNA gene typing and produced new serotyping monoclonal antibodies (MAbs) to characterize the C:NST:NSST isolates. We were able to select two hybridoma cells that recognizes PorB and PorA proteins. Among the 255 strains initially characterized as NST:NSST, 75% (n=191) of them became completely serotyped as 23:P1 .14-6. Sy 16S RNA ribossomal typing, these strains showed the same pattern suggesting strain clonality. Our data demonstrate the introduction of a new clone of Neisseria rneningitidis C presenting the phenotype C:23:P1.14-6 and that is being responsible for the increase of the cases of DM caused by this serogroup in Great Sao Paulo.

Meningococcal meningitis is a burden in the African meningitis belt. Before 2010, Neisseria meningitidis serogroup A (N. meningitidis A) was the predominant pathogen causing deathly epidemics. MenAfriVac® vaccine protects against N. meningitidis A. It was introduced in 2010 into highest meningitis risk health districts. There was limited data on the effects of MenAfriVac®, mainly on the degree of relationship between N. meningitidis A and the MenAfriVac® immunization. The social ecological model was used as a theoretical framework for this study. The purpose of this quantitative study was to assess the effectiveness of MenAfriVac® from 2010 to 2017 in 21 out of 26 countries of the African meningitis belt. The four research questions contributed to establishing the effects of MenAfriVac®. An interrupted time series design and nonprobability sampling were used. Secondary data were retrieved from World Health Organization database. The binomial negative regression and Pearson’s Chi-Square tests were used. The study found that after the MenAfriVac® introduction there were 39% decline of incidence rate of the meningitis suspected cases (IRR 0.61, 95% CI 0.48 – 0.79, p < .001), a high degree of relationship between N. meningitidis A and MenAfriVac® immunization (χ2 (1) = 11039.49, p = 0.000, Phi = 0.657, P=0.000), 99% decline of the risk of N. meningitidis A (RR 0.01, 95% CI 0.08-0.013), and 99.6% decline of risk of epidemic due to N. meningitidis A (RR 0.004, 95% CI 0.001-0.016). The study demonstrated that high MenAfriVac® coverage and enhanced surveillance are pivotal to reduce the meningitis burden. Results will be used to inform policy and public health practice to reduce the meningitis cases and improve quality of live in the community.

Padronizou-se a PCR-Multiplex para a detecção do DNA de Neisseria meningitidis. Para tanto os primers escolhidos foram: RW01, DG74 e COR28 baseados na subunidade menor do ribossomo (16S rRNA) que apresenta regiões de seqüências conservadas encontradas em todas as bactérias conhecidas. Os primers RW01 e DG74 amplificaram o fragmento universal bacteriano de 370 bp e, os primers RW01 e COR28, o fragmento específico de N. meningitidis de 279 bp em uma única etapa. Os resultados obtidos nas amostras de LCR de 168 pacientes pelos métodos de cultura e PCR-Multiplex quando comparados à bacterioscopia mostraram que tal técnica apresentou alta sensibilidade (91,3%) no estudo de amostras de LCR de bacterioscopia positiva, enquanto que a cultura apresentou resultados menores (19,7%). Nas amostras de LCR com bacterioscopia negativa a sensibilidade da PCR-Multiplex (57,8%) também foi mais elevada do que da cultura (10%). Estes dados sugerem que a técnica aqui padronizada é altamente promissora para ser utilizada como método diagnóstico da meningite meningocócica, especialmente nos casos de pacientes submetidos à terapia antibiótica prévia.
The PCR-multiplex technique was standardized to detect N.meningitidis DNA. It was used universal primer for all bacteria showing sequence of minor subunit of 16S ribossome regions (RW01, DG74) by amplification of 370bp fragment and another (COR28) for specific sequence of N. meningitidis, amplifying 279bp fragment in one step. The results obtained in CSF samples of 168 patients by culture and PCR-Multiplex technique when compared with microscopy showed high sensitivity (91,3%) in samples with positive microscopy (81) to Gram negative diplococcus, however the culture presented only 19, 7% of positivity in the same samples. In other hand the CSF samples with negative bacterioscopy (67) the PCR-Multiplex sensitivity (57,8%) was higher to culture (10,0%) too. These data indicate a high sensitivity and specificity of PCR as a tool for a rapid diagnosis of meningococcal meningitis, mainly in that patient submitted to previous antibiotic therapy as in case of this work (90% of patients) besides the possibility of a rational practice of specific treatment.

E realizado o estudo epidemiológico das meningites de etiologia indeterminada no Municlpio de São Paulo, no período de 1960 a 1977. O trabalho é apresentado em quatro partes. Na primeira parte são apresentados e discutidos os diferentes modos de diagnóstico etiológico das meningites (presuntivos e de certeza) e é enfatizada a necessidade do estudo das meningites indeterminadas. Na segunda parte é apresentado o comportamento epidemiológico da doença meningocócica no Município de São Paulo, no período 1960 a 1977. Esta apresentação é feita visando comparar e procurando verificar a influência da doença meningocócica no comportamento epidemiológico das meningites de etiologia indeterminada. Na terceira e quarta partes são apresentados os caracteres epidemiológicos das meningites de etiologia indeterminada. Inicialmente os casos são classificados em meningites indeterminadas de provavel etiologia bacteriana ou viral, utilizando-se como parâmetro para a classificação o percentual de neutrófilos no liquor. Em seguida são apresentados e analisados os comportamentos segundo variaveis da pessoa (idade e sexo), do local de residência (segundo distritos e areas homogêneas do município), do tempo (variação anual e mensal), modo diagnóstico, evolução clinica, tempo de hospitalização. O estudo mostra que entre 1972 e 1977 ocorreu no Município de São Paulo um aumento significativo de casos de meningite de etiologia indeterminada; grande parte dos casos, provavelmente, era constituída por casos de meningite meningocócica, dos quais não foi possivel a determinação etiológica. Ocorreu também, concomitantemente, um aumento significativo de meningites de provável etiologia viral. O percentual de neutrófilos no liquor (primeiro exame) em épocas epidêmicas, pode ser utilizado como parâmetro para classificacão epidemiológica meningites segundo etiologia provavelmente bacteriana ou viral.
The epidemiological behaviour of the meningitis of undetermined etiology within the Municipality of São Paulo, during the period from 1860 to 1977, is studied. The work consists of four parts. In the first part, the different forms of etiological diagnosis of meningitis (either presumptives ar certain) are submitted and discussed, as well as the need study of the undetermined meningitis. In the second part the epidemiological behaviour of the meningococcal diseases within the Municipality of São Paulo, during the period from 1960 to 1977, is submitted. Such a presentation is made with the purpose of comparison, and trying to verify the influence of the meningococcal disease on the epidemiological behaviour of the meningitis of undetermined etiology. In the third and fourth parts, the epidemiologic characters of the meningitis of undetermined etiology are submitted. At first, the cases are classified as undetermined meningitis of probable bacterial or viral etiology, using as parameter for the classification the percentile number of neutrophiles in the liquor (first examination). Then, there are submitted and analysed the behaviour as per the variables of the person (age and sex), place of residence (according to districts and homogeneous areas within the Municipality), time (annual and monthly variation), form of diagnosis, clinic evolution, and hospitalization period. It is concluded that between 1972 and 1977 there was in the Municipality of São Paulo, an epidemics of meningitis of undetermined etiology; most of the cases were, probably, constituted by cases of meningococcic meningitis, of which it was not possible to make the etiologic determination. There was, also, al the same time, an important increase of diagnosis of meningitis of probable viral etiology. The percentile number of neutrophiles in the Liquor (first examination), in epidemic times, may be used as an epidemiological parameter for the classification of the meningitis, as per the etiology, probably bacterial or viral.

Meningococcal meningitis is a major public health problem in an area of sub-Saharan Africa known as the meningitis belt. In this region, thousands of people fall ill each year during highly seasonal meningitis outbreaks. At frequent but unpredictable times, large epidemics of meningitis sweep the belt. Despite the gravity of the problem, much is still unknown about what drives this pattern of epidemics. The aim of this thesis is to use mathematical models to gain insights into the epidemiology of meningococcal meningitis in the meningitis belt. Firstly, several simple deterministic models are analysed. They highlight the importance of immunity following both asymptomatic carriage and invasive meningitis. For a broad range of plausible parameter values, seasonal variation of the rate of meningococcal transmission results at long, irregular intervals. Realistic sized repeated epidemics are not possible if only the ration of cases to carriers changes seasonally.

The primary approach to the control of meningococcal disease remains effective vaccination programmes in susceptible populations. Vaccines against serogroups A, C, W and Y offer broad protection against meningococci and both polysaccharide and conjugate quadrivalent vaccines are licensed for use in the UK. Previous studies have assessed the antibody response to meningococcal polysaccharide and conjugate vaccines, but there is limited information on the nature of the B cell response to these antigens. As part of a clinical trial using both polysaccharide (MenACWY-PS) and conjugate (MenACWY-CRM) vaccines in adult volunteers, this DPhil reports the analysis of subsets of antigen specific B-cells produced in response to either vaccine. Prior MenACWY-PS impaired the response to a subsequent dose of MenACWY-CRM. This may be due to MenACWY-PS driving terminal differentiation of antigen specific cells into plasma cells, without replenishment of the memory B cell pool. In addition, despite prior data indicating that it may act as a thymus dependent antigen, the serogroup A polysaccharide component of MenACWY-PS appears to behave in the same way as serogroup C, W & Y polysaccharide components. Antibody molecules recognise and bind to a multitude of conformational epitopes. This variability is enabled by the complexities of immunoglobulin variable domain gene recombination which can generate a vast potential repertoire of unique antibody molecules. However, the diversity of the antibody repertoire is more restricted against specific antigens and within defined B cell subsets. In this DPhil, ‘next generation’ sequencing technologies were used to investigate the diversity of the B cell variable domain before and after vaccination of adult volunteers. Individuals at baseline were found to have distinct antibody repertoires. Vaccination with a Haemophilus influenzae type b (Hib) conjugate vaccine resulted in an oligoclonal antibody response, with enrichment for Hib specific canonical antibody sequences.

O presente trabalho estuda algumas características epidemiológicas de 241 casos de Meningite tuberculosa de pessoas residentes na Grande São Paulo, nos anos de 1982 e 1983. O levantamento de casos foi realizado no Centro de Investigações de Saúde e outras fontes oficiais de informação e complementado pela visitação domiciliária que representou um recurso inestimável para o esclarecimento dos dados. Os casos foram analisados por região de residência, aspectos individuais, núcleos familiares, história da doença, hospitalização, seqüência de tratamento e conhecimento sobre a doença. Os resultados identificam condições insatisfatórias de vida na maioria da população, demora no diagnóstico por falhas assistenciais, alta letalidade hospitalar e desconhecimento do modo de transmissão e prevenção da tuberculose pela maioria das pessoas entrevistadas. O grupo de menores de 5 anos de idade foi o mais comprometido pela ocorrência de seqüelas e, a maior letalidade oi na faixa de 7-12 meses. Ao final do estudo, houve 45,7 por cento de cura, 27,8 por cento de óbito, 13,3 por cento de abandono. Em 13,2 por cento dos casos, alguns permaneciam em observação e outros desconhecidos pelo sistema de controle de notificações.
The present paper studies some epidemiological characteristics of 241 cases of tuberculous miningitis in persons living in the Great São Paulo, State of São Paulo, Brazil, in 1982 and 1983. The survey of cases was worked out at the Center of Health Investigation; other official sources have been consulted, too, being work complemented by domiciliary visitings which represented an invaluable resource for data enlightenment. The cases were analysed taking into account region of dwelling, individual aspects, familiar nucleus, disease history, hospitalization, treatment follow-up and knowledge about the disease. Results identify unsatisfactory life conditions for the majority of the population under study; delay in diagnosing the disease due to failures in assistance; high rate of hospital lethality and lack of knowledge on how tuberculosis is transmitted and prevented by the majority of the persons interviewed with. The age group of children below five years was the one most implicated in as to the occurence of the highest rate of lethality was presented by children aged 7-12 months. At the end of the study, there were 45.7 per cent of healings; 27.8 per cent of deaths; 13.3 per cent of treatment abandonment. In 13.2 per cent of the cases, some persons continued under observation and others remained unknown by the Health System of notifications.

Streptococcus pneumoniae (SP) meningitis remains a significant cause of global childhood mortality, and a potential epidemic pathogen despite improving vaccinations schemes. Neuropathology in pneumococcal meningitis (PM) is attributed to both bacterial products and the host-mediated proinflammatory response. Proinflammatory cytokine interferon-gamma (IFNγ) is associated with macrophage activation and regulating MHC Class I expression in innate immunity. IFNγ gene knockout (GKO) mice inoculated with ~5 x 103 CFU of SP/mouse were significantly protected against fatal PM, compared to C57Bl/6 WT mice, in both serotype 3 WU2 and serotype 4 TIGR4 strains. TIGR4 PM was comparatively more virulent than WU2 PM. In murine PM, IFNγ regulated induction of chemokines MCP1, CXCL9, CXCL10, the IRGM1, IRGM3 proteins and nitric oxide synthase 2 (NOS2). In WU2 PM, production of IFNγ was attributed to infiltrating Natural Killer (NK) cells, and dependent upon activation of the inflammasome complex and interleukin-18. NOS2 GKO exhibited a significantly improved survival phenotype, and reduced blood brain barrier dysfunction, compared to WT mice. Intracellular staining revealed Ly6Chi monocytes and NK cells as the source of IFNγ-dependent NOS2 in PM. Overall, this thesis addresses IFNγ production and its contribution to experimental PM pathology.

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Descreve um estudo de Coorte Descritivo Retrospectivo, realizado com dados obtidos através de prontuários de pacientes internados no Instituto Estadual de Infectologia Säo Sebastiäo, no período de 11/96 a 06/97, que tiveram diagnóstico definitivo de meningite bacteriana ou viral. Procedeu-se às estimativas das gravidade e letalidade, de ambas as meningites, comparando-as entre si, bem como dos respectivos agentes etiológicos específicos. Com base nos 204 pacientes, 141 dos quais, portadores de meningite bacteriana e 63, de meningite viral, viu-se que as primeiras se apresentaram mais graves e mais letais que estas últimas, com excessos de risco de 17,6 e 7,8 por cento, respectivamente. Também evidenciou-se que, apesar de todas as infecçöes bacterianas apresentarem casos graves e incidência de óbitos, a que teve maior número destes desfechos foi a meningite pneumocócica, enquanto que, dentre as virais, a meningite por Herpes simples vírus, foi a única a apresentar tais eventos. Os maiores preditores para a gravidade foram a meningite pneumocócica, a meningite por Herpes simples vírus e a idade de 15 anos ou mais. Já para a letalidade, os preditores, além destes patógenos, foram os menores de 1 ano e evoluçäo clínica para o coma, na meningite bacteriana, e a idade de 15 anos ou mais e evoluçäo para torpor ou coma na viral. Ainda constatou-se que as características liquóricas seguem um padräo bem definido para cada uma das meningites em estudo.
This is a Retrospective Descriptive Cohort study, accomplished through reference book, with patients interned at the São Sebastião State Institute of Infectology, in Rio de Janeiro City, Brazil, in the period from 11/96 to 06/97, with definitive diagnosis of bacterial or viral meningitis. It was proceeded to the estimates of the severity and mortality, of both meningitis, comparing them to each other, as well as the respective pathogens. Based on the 204 patients, 141 of the which, carriers of bacterial meningitis and 63, of viral meningitis, the first ones came more severe and more lethal than these last ones, with excesses of risk of 17,6 and 7,8%, respectively. It was also evidenced that, in spite of all the bacterial infections they present severe cases and obits incidence, the one that had larger number of these was the pneumococcal meningitis, while, of the viral ones, the meningitis for Herpes simplex virus, was the only to present such events. The most importants predictores for the severity were pneumococcal meningitis, herpes simplex virus meningitis and the 15 years-old age or more. Already for the mortality, the predictores, besides these pathogens, was last then 1 year old and clinical evolution for the coma, in the bacterial meningitis, and the 15 years-old age or more and evolution for torpor or coma, in the viral one. It was still verified that the cerebrospinal fluid (CSF) characteristic follows a pattern very defined for each one of the meningitis in study.

Philosophiae Doctor - PhD
Neisseria meningitidis (the meningococcus), the causative agent of meningococcal disease (MD) was identified in 1887 and despite effective antibiotics and partially effective vaccines, Neisseria meningitidis (N. meningitidis) is the leading cause worldwide of meningitis and rapidly fatal sepsis usually in otherwise healthy individuals. Over 500 000 meningococcal cases occur every year. These numbers have made bacterial meningitis a top ten infectious cause of death worldwide. MD primarily affects children under 5 years of age, although in epidemic outbreaks there is a shift in disease to older children, adolescents and adults. MD is also associated with marked morbidity including limb loss, hearing loss, cognitive dysfunction, visual impairment, educational difficulties, developmental delays, motor nerve deficits, seizure disorders and behavioural problems. Antimicrobial peptides (AMPs) are molecules that provide protection against environmental pathogens, acting against a large number of microorganisms, including bacteria, fungi, yeast and virus. AMPs production is a major component of innate immunity against infection. The chemical properties of AMPs allow them to insert into the anionic cell wall and phospholipid membranes of microorganisms or bind to the bacteria making it easily detectable for diagnostic purposes. AMPs can be exploited for the generation of novel antibiotics, as biomarkers in the diagnosis of inflammatory conditions, for the manipulation of the inflammatory process, wound healing, autoimmunity and in the combat of tumour cells. Due to the severity of meningitis, early detection and identification of the strain of N. meningitidis is vital. Rapid and accurate diagnosis is essential for optimal management of patients and a major problem for MD is its diagnostic difficulties and experts conclude that with an early intervention the patient’ prognosis will be much improved. It is becoming increasingly difficult to confirm the diagnosis of meningococcal infection by conventional methods. Although polymerase chain reaction (PCR) has the potential advantage of providing more rapid confirmation of the presence of the bacterium than culturing, it is still time consuming as well as costly. Introduction of AMPs to bind to N. meningitidis receptors could provide a less costly and time consuming solution to the current diagnostic problems. World Health Organization (WHO) meningococcal meningitis program activities encourage laboratory strengthening to ensure prompt and accurate diagnosis to rapidly confirm the presence of MD. This study aimed to identify a list of putative AMPs showing antibacterial activity to N. meningitidis to be used as ligands against receptors uniquely expressed by the bacterium and for the identified AMPs to be used in a Lateral Flow Device (LFD) for the rapid and accurate diagnosis of MD.

Background Cryptococcal meningitis and tuberculosis are the lead causes of mortality in late stage HIV. I examine the treatment and prevention of Cryptococcal disease and early detection of Tuberculosis in Sub- Saharan Africa. Methods I conducted 4 studies: 1. Dose-response effect of high dose fluconazole for HIV-associated Cryptococcal Meningitis. 2. Cryptococcal Antigen Screening in Patients Initiating ART: A Prospective Cohort Study 3. Evaluation of a Public-sector, Provider-initiated Cryptococcal Antigen Screening and Treatment Program. 4. The predictive value of urine lipoarabinomannan lateral-flow assay for incident tuberculosis, hospitalisation and / or death in outpatients during their first year of ART. Results The early fungicidal activity of Fluconazole was significantly greater for 1200mg than it was for 800mg. Incident CM was associated with higher plasma/serum LFA titers. 12-month mortality was 25% in CrAg-positive patients compared with 11.5% in CrAg-negative patients despite pre-emptive fluconazole treatment. 4 out of 10 serum CrAg positive patients agreeing to an LP were found to have evidence of meningeal infection. In the provider initiated CrAg screening program in the public health sector only 26.6% of eligible patients were screened during the first year of the program. Patients testing LAM-positive had a median time to TB treatment of 7 days and were more likely to require hospitalisation and die within the first year of ART. Conclusion Where Amphotericin B is not available or safe to administer, 1200mg of Fluconazole clears infection more rapidly than 800mg. CrAg screening and pre-emptive treatment with fluconazole in patients with late stage HIV reduced the number of cases of CM compared to historic controls. Further studies are needed to refine the treatment of CrAg positive patients. Provider initiated CrAg screening did not achieve optimal levels and may have caused delay in ART initiation. Screening for urinary LAM may reduce the time to TB treatment by 1 week and allow early identification of patients at the highest risk of adverse events in the first year of ART.

Master of Public Health - MPH
Meningitis is a common opportunistic infection and an important cause of mortality among people living with HIV and AIDS globally. This study investigated meningitis in adults living with HIV and AIDS admitted to the medical wards of Livingstone tertiary hospital in Port Elizabeth in 2018 and determined the prevalence of its aetiological types, clinical presentations, diagnostic challenges, treatment outcomes and predictors of prognosis.

Orientador: Marcelo Lancellotti
Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia
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Resumo: Vesículas da membrana externa ou outer membrane vesicles (OMV) são nanopartículas liberadas no meio de cultura durante o crescimento bacteriano resultantes de evaginações da membrana externa. A OMV é composta por antígenos presentes na membrana externa bacteriana, como porinas e lipo-oligossacarídeos - LOS, proporcionando a estas estruturas um efeito imuno-estimulatório, sendo alvo potencial para a produção de vacinas. Este trabalho teve como objetivo analisar a utilização de OMV de Neisseria meningitidis IAL 2443, linhagem pertencente ao sorogrupo B causadora de meningites no estado de São Paulo, com nanopartículas de sílica mesoporosa SBA-15 e SBA-16 no processo de imunização vacinal. Utilizou-se o processo fermentativo em estado semi-sólido para o crescimento bacteriano e um processo de ultracentrifugação para isolamento da OMV. Comparou-se a produção de OMV de linhagens de N. meningitidis selvagens IAL 2443 e C2135 e mutante knock-out para o gene responsável pela produção de pilina - C2135_pilE. A atividade imunológica foi verificada por meio da imunização em camundongos Swiss e a verificação da citotoxicidade foi realizada em cultura celular utilizando células VERO e NHI 3T3. Os resultados mostraram que as diferentes linhagens de N. meningitidis possuem cinéticas de produção de OMVs distintas em tempo e quantidade. A não expressão de pilE também afeta a cinética da produção dessas estruturas. O uso de adjuvantes de sílica mesoporosa SBA- 15 e SBA- 16 com a OMV IAL 2443 aumenta o reconhecimento pelo sistema imunológico, consequentemente elevando o número de anticorpos específicos frente a mesma cepa bacteriana e a outras linhagens do mesmo sorotipo e sorotipos diferentes. O teste de citotoxicidade em células VERO e NHI 3T3 demonstrou inocuidade nas preparações com SBA- 15 e SBA- 16, atestando a segurança no uso destas preparações como adjuvantes vacinais. O estudo demonstrou que a metodologia utilizada para produção de OMV é vantajosa do ponto de vista quantitativo e econômico e o uso de SBA- 15 e SBA- 16 apresenta potencial como adjuvante em vacinas baseadas em OMV
Abstract: Outer membrane vesicles or OMV are nanoparticles released into the culture medium during bacterial growth resulting from evaginations of the outer membrane. The OMV consists of antigens present in the bacterial outer membrane, like porins and lipooligosaccharides - LOS, these structures provide an immuno-stimulatory effect, and potential target for vaccine production. This study aimed to analyze the use OMV of Neisseria meningitidis IAL2443, belonging to serogroup B strain causing meningitis in São Paulo, as vaccine using the mesoporous silica SBA-15 and SBA-16 in the immunization process. Fermentation process was used in semi-solid state for bacterial growth and a process of ultracentrifugation for the isolation these OMV. It was compared to OMV production of strains of N.meningitidis strains IAL 2443 and C2135, and knock-out mutant for the gene responsible for producing pilin - C2135_pilE. The immunological activity was measured by immunization in Swiss mice and verification of cytotoxicity was performed in cell culture using VERO cells and NIH 3T3. The results showed that different strains of N. meningitidis have OMVs kinetics of production of different time and quantity. The expression does not pile also affects the kinetics of production of these structures. The use of adjuvants mesoporous silica SBA-15 and SBA-16 with the IAL2443 OMV increases the recognition by the immune system, thus bringing the number of specific antibodies against the same bacterial strain and other strains of the same serotype and different serotypes. The cytotoxicity test in NIH 3T3 cells showed VERO and safety in preparation adjuvanted with SBA-15 and SBA-16, confirming the safe use of these preparations as vaccine adjuvants. The study showed that the methodology used for the production of OMV is advantageous from the point of view of quantity and cost and use of SBA-15 and SBA-16 has potential as an adjuvant in vaccines based on OMV
Mestrado
Bioquimica
Mestre em Biologia Funcional e Molecular

Tuberculosis, caused by the bacterium Mycobacterium tuberculosis, kills 1.4 million people each year. Tuberculous meningitis (TBM) is the most severe manifestation of disease caused by M.tuberculosis with high mortality, and disabling neurological sequelae common among those who survive. Early diagnosis and initiation of antituberculous chemotherapy are the strongest prognostic factors for a successful outcome, yet diagnosis of TBM remains challenging, with conventional microbiological techniques for detection of M.tuberculosis either slow or insensitive. Standardised treatment regimens use the same drugs and dosages as pulmonary TB, despite the variability in CSF penetration of these drugs. Treatment regimens for TBM have never been optimized through randomized controlled trials and little is known about the long-term outcomes of those who survive. In addition children with TBM may require different dosages to adults.

Orientador: Marcelo Lancellotti
Dissertação (mestrado) - Universidade Estadual de Campinas, Insituto de Biologia
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Resumo: Neisseria meningitdis, ou meningococo, é uma bactéria comensal da nasofaringe humana. Contudo, algumas linhagens meningocócicas ocasionalmente ultrapassam a mucosa respiratória e a barreira hematoencefálica e causam enfermidades como meningite e septicemia. Como a espécie humana é o único hospedeiro natural para esse patógeno, estudos in vitro com modelos celulares são uma ferramenta importante para a análise da interação entre o meningococo e seu hospedeiro. Este trabalho teve por objetivo avaliar a influência de diferentes linhagens de N. meningitidis na adesão celular, morfologia e expressão de quimiocinas inflamatórias em culturas in vitro de células humanas. Tais parâmetros também foram avaliados em um sistema in vitro de co-cultura celular entre células de origem nervosa e endotelial a fim de mimetizar a barrreira hematoencefálica humana. Os resultados obtidos indicam que a adesão de diferentes linhagens bacterianas em células humanas de sítios específicos do processo infeccioso do meningococo, como Hep-2 (laringe), NCIH460 (pulmão), Hec1b (endotelial) e NG97 (neuroglia) foi capaz de mimetizar o processo fisiopatológico deste microrganismo. Em condições in vitro, células de origem nervosa mostraram-se mais suscetíveis à infecção nos parâmetros avaliados como uma elevada expressão de TNF-?, quimiocina característica em infecções meningocócicas. A linhagem B4 destacou-se entre as linhagens meningocócicas estudadas, apresentando elevados percentuais de adesão em células humanas com valor máximo de adesão em NG97. Células HEp-2 apresentaram poucas alterações morfológicas significativas frente à infecção por N. meningitidis. Tais resultados podem estar associados ao fato do trato respiratório superior ser o habitat natural do meningococo, no qual a interação entre este patógeno e as células hospedeiras seja comensal e não-invasiva. O modelo mimético de barreira hematoencefálica, realizado em transwell, indicou comunicação entre as células que o compõem e uma maior expressão dos níveis de quimiocinas inflamatórias quando comparada à infecção desta bactéria por cada uma das células estudadas isoladamente. Tal modelo foi capaz de mimetizar a barreira hematoencefálica, fato o que torna possível sua aplicação em estudos da passagem de fármacos e outros patógenos por essa barreira.
Abstract: Neisseria meningitidis,or meningococci, is a commensal bacterium of the human nasopharynx. However, occasionally some meningococcal strains can cross the respiratory mucosa and the blood-brain barrier and cause life-threatening diseases such as meningitis and septicaemia. Since the human species is the only natural host for this pathogen, in vitro studies with cellular models are an important tool for the analysis of the meningococci and its host. The aim of this present work was to value the influence of several strains of N. meningitidis in cellular adhesion, morphology and inflammatory chemokines expression in human cells cultivated in vitro. These parameters were also evaluated in a co-cultivated cellular system with glial and endothelial cells aiming mimicking the human blood brain barrier. The results indicate that the adhesion of different bacterial strains from specific sites of infectious process of meningococci as Hep-2 (larynges), NCIH460 (lung), Hec1b (endothelial) and NG97 (glial cells) was capable of mimicking, partially, the pathophysiology of this microorganism. In in vitro conditions, cells from nervous origin showed more susceptibility to infection then others in this evaluated parameters such as a high TNF-? expression, a common chemokine in meningococcals diseases. The B4 strain distinguished from the others by presenting high rates of adhesion in human cells with maximum value on NG97. HEp-2 cells showed few expressives morphologic alterations after meningococcal infection. These results may be associated with the fact that the upper human respiratory tract is the meningoccci natural habitat, in which the interaction between this pathogen and host cells are commensal and not-invasive.The mimicking blood-brain barrier model, performed in transweel, has demonstrated some communication between the cells and greater expression of inflammatory chemokines when compared to the infection in isolated cells. This model was capable of mimicing the blood-brain barrier, which makes its application possible in studies of drugs and others pathogens who might crossover though this barrier.
Mestrado
Bioquimica
Mestre em Biologia Funcional e Molecular

Experimental meningitis was induced in pigmented guinea pigs by subarachnoid inoculation of \(1 \times 10^9\) Escherichia coli K-12 or \(3 \times 10^7\) CFU Streptococcus pneumoniae serotype 2 D39 (NCTC 7466) or PLN-A, \(\Delta\)NA1 or \(\Delta\)HY1, defined isogenic derivatives of D39 deficient in pneumolysin, neuraminidase or hyaluronidase respectively. All animals developed a meningeal inflammatory response and a labyrinthitis. Hearing loss in pneumococcal meningitis was measured by recording the evoked auditory nerve compound action potential from the round window membrane. Animals infected with PLN-A sustained significantly less hearing loss than those infected with wild-type D39 (12 dB vs. 50 dB 12 h post inoculation; P<0.0001), Neuraminidase deficiency did not alter the course of the meningeal inflammatory response nor affect hearing loss. The \(\Delta\)HY1 mutant survived poorly in the cerebrospinal fluid and blood but still caused hearing loss. Both pneumococcal and E. coli meningitis induced specific ultrastructural lesions in the organ of Corti as judged by high-resolution scanning and transmission electron microscopy, and these lesions were most severe with pneumolysin-sufficient pneumococcal infection. Microperfusion of \(5\times10^6\) CFU S.pneumoniae D39 directly into the scala tympani of guinea pigs also resulted in electrophysiological and ultrastructural damage to the organ of Corti that could be diminished by pretreatment with antibiotics. The data confirm the cochlea as the site of meningogenic deafness. They suggest that pneumolysin expression is chiefly responsible for meningogenic deafness and that if pneumococci invade the inner ear during bacterial meningitis, cochlear deafhess will rapidly ensue.

Includes bibliographical references.
[Background] Tuberculous meningitis (TBM) is a common form of tuberculosis in high TB incidence settings. However, the burden of disease and outcome in affected adults is unknown in Cape Town. The diagnosis of TBM is often challenging, particularly in HIV co-infected patients and no standardized clinical case definition exists. An emerging complication that contributes to poor outcome in HIV-associated TBM is neurological TB immune reconstitution inflammatory syndrome (TB-IRIS). [Methods] A consensus clinical TBM case definition was developed following a TBM meeting that I co-ordinated. I led two observational studies that determined the burden of HIV-associated TBM and neurological TB-IRIS at a district-level hospital in Cape Town. Patients with HIV-associated TBM were prospectively enrolled in a third cohort study to determine the clinical and immunological characteristics of paradoxical TBM-IRIS. [Results] TBM accounted for 57% of meningitis cases over a 6-months period; 88% of these patients were HIV-infected. At six months follow-up, mortality in HIV-associated TBM patients was 48%. Neurological TB-IRIS accounted for 21% of patients who presented with central nervous system (CNS) deterioration during the first year of antiretroviral therapy (ART) over a one-year period. TBM-IRIS developed in 47% of HIV-associated TBM patients and associated with extensive cerebrospinal fluid (CSF) inflammation both at TBM diagnosis and at TBM-IRIS presentation. Patients who did not develop TBM-IRIS, but who were culture-positive for Mycobacterium tuberculosis from CSF at TBM diagnosis, showed an immunological phenotype similar to TBM-IRIS patients; however neutrophils were increased in TBM-IRIS patients compared to culture-positive TBM-non-IRIS patients, both at TBM diagnosis and two weeks after ART initiation. [Conclusions] HIV-associated TBM is a common cause of meningitis with a poor outcome in Cape Town. TBM-IRIS is a frequent complication of ART in HIV-associated TBM patients. CSF Mycobacterium tuberculosis culture positivity drives an inflammatory response that manifests as TBM-IRIS in most, but not all TBM patients. Neutrophils associate closely with the CNS inflammation that characterizes TBM-IRIS. An intensified TB treatment regimen with increased CSF penetration early during TB treatment may lead to improved mycobacterial clearance from the CNS, which may result in improved outcome during TBM treatment and a reduced frequency of TBM-IRIS. We aim to test this hypothesis in future studies.

A model for meningitis is developed by adding a class of carriers to the basic SIR model. This model is used to analyze the impact a vaccination program can have on the health of the population of epidemic prone countries. Analysis of the model shows the local stability of the disease free equilibrium, the existence of an endemic equilibrium and computation of the reproduction number, ℜ0 . Using a MATLAB program we simulate a time course of the model using parameters gathered from the World Health Organization. The numerical solution demonstrates that our reproduction number was correct. We thenconcluded that a high infection transmission rate requires a high vaccine rate.

Thesis (PhD)--Stellenbosch University, 2015.
ENGLISH ABSTRACT: Tuberculous meningitis (TBM) continues to be an important cause of mortality and neurological disability in resource-limited countries. Many questions remain about the best approaches to prevent, diagnose, and treat TBM, and there are still too fewanswers. The aim of this dissertation was to challenge current management strategies in childhood TBM. Accurate prediction of outcome in TBM is of critical importance when assessing the efficacy of different interventions. I conducted a retrospective cohort study of 554 children with TBM less than 13 years of age admitted to Tygerberg Children’s Hospital over a 20 year period (1985-2005) and reclassified all patients according to the criteria of all the currently available staging systems in childhood TBM (chapter 4). In this study, I found that the “Refined Medical Research Council (MRC) staging system after 1 week” had the highest predictive value of all TBM staging systems. It is created by subdivision of stage 2 (2a and 2b) of the existing MRC staging system. Additionally, I proposed and validated a simplified TBM staging system which is less dependent on clinical ability and neurological expertise than current staging systems. The simplified staging system was termed the “Tygerberg Children’s Hospital Scale” (TCH) and relies solely on the patient’s ability to visually fixate and follow and the motor response to pain on both sides. It demonstrated excellent predictive power of outcome after 1 week and did not differ significantly from the “Refined MRC staging system” in this regard. The optimal anti-TB drug regimen and duration of treatment for TBM is unknown. It has been suggested that intensive short-course (6 months) anti-TB therapy may be sufficient and safe. I conducted a prospective descriptive study of 184 consecutively treated children with TBM and found that short-course intensified anti-TB therapy aimed at treating TBM patients (anti-TBM therapy) is sufficient and safe in both HIV-uninfected and HIVinfected children with drug susceptible TBM (chapter 5). The overall study mortality of 3.8% at completion of treatment compares favourably with the median mortality rate of 33% (range 5-65%) reported in a recent review describing outcome in TBM treatmentstudies. TB-immune reconstitution inflammatory syndrome (IRIS) is a potentially life-threatening complication in HIV-infected children with TB of the central nervous system. Little is known about the incidence, case fatality, underlying immunopathology and treatment approaches in HIV-infected children with neurological TB-IRIS. In a case series, I found that neurological TB-IRIS should be considered when new neurological signs develop after initiation of antiretroviral therapy (ART) in children with TBM (chapter 6.1). Manifestations of neurological TB-IRIS include headache, seizures, meningeal irritation, a decreased level of consciousness, ataxia and focal motor deficit. I also discussed the rational for using certain treatment modalities, includingthalidomide. Neurological tuberculous mass lesions (tuberculomas and pseudo-abscesses) may develop or enlarge in children on anti-TBM treatment. These lesions respond poorly to therapy, and may require surgical excision, but may be responsive to thalidomide, a potent inhibitor of tumour necrosis factor-alpha (TNF-alpha). The optimal dose and duration of thalidomide therapy and the correlation with magnetic resonance imaging (MRI) is yet to be explored. The primary objective of our next study was to investigate whether serial MRI is useful in evaluating treatment response and duration of thalidomide therapy (chapter 6.2). A secondary objective was to determine the value of thalidomide in the treatment of these lesions. In a prospective observational study over three years, serial MRI was performed in 16 consecutive children compromised by TB pseudo-abscesses who were treated with thalidomide. The rapid clinical response of most patients suggests that thalidomide provides substantial clinical benefit in this clinical context. I also identified a MRI marker of cure that is evolution of lesions from early stage “T2 bright” with edema to “T2 black.” This finding could be useful in the future management of these patients. Transcranial Doppler imaging (TCDI) is potentially a valuable investigational tool in children with TBM, a condition often complicated by pathology relevant to Doppler imaging such as raised intracranial pressure (ICP) and cerebral vasculopathies. Serial TCDI was performed on 20 TBM children with the aim of investigating cerebral haemodynamics and the relationship between pulsatility index (PI) and ICP (chapter 6.3). In this study, I found that TCDI-derived pulsatility index (PI) is not a reliable indicator of raised ICP in children with tuberculous hydrocephalus which I attributed this to individual variation of tuberculous vascular disease, possibly compromising cerebral vascular compliance and resistance. The study did confirm the efficacy of medical therapy in children with tuberculous communicating hydrocephalus. In all cases, the ICP normalized within 7 days after initiation of acetazolamide and furosemide. In the same cohort of children with TBM I also measured cerebral blood flow velocities (BFV) in the anterior cerebral artery (ACA), middle cerebral artery (MCA) and posterior cerebral artery (PCA) on admission and after day 3 and 7. I found persistent high BFV in all the basal cerebral arteries suggesting stenosis due to vasculitis rather than functional vasospasm. Additionally, I found that complete MCA occlusion, subnormal mean MCA velocities (less than 40 cm/s) and a reduced PI (less than 0.4) correlated with radiological proven large cerebral infarcts. No side-to-side differences in MCA BFV or subnormal PI’s were detected in four TBM children with territory infarcts on admission. I attributed this to the occlusion of a limited number (one or two) of the 9 MCA perforators which has been shown not to affect the hemodynamics of the MCA. I concluded by highlighting the many questions that remain about the best approaches to prevent, diagnose, and treat TBM (chapter 2). In a second literature review, aimed at clinicians working in resource-limited countries, I describe novel approaches to the management of childhood TBM, including a treatment algorithm for tuberculous hydrocephalus, the role for short-course intensified anti-TBM treatment and home-based anti-TBM treatment (chapter 3). Even with the best diagnostic and treatment modalities, outcome in childhood TBM will remain poor if diagnosis is delayed. Our efforts should be on increased awareness and earlier diagnosis.
AFRIKAANSE OPSOMMING: Tuberkuleuse meningitis (TBM) bly ‘n belangrike oorsaak van mortaliteit en neurologiese ongeskiktheid in lande met beperkte hulpbronne. Baie vrae oor die beste benaderings tot voorkoming, diagnose en behandeling van TBM bly bestaan en daar is steeds te min antwoorde. Die doel van die verhandeling was om huidige behandelingstrategieë van tuberkuleuse meningitis (TBM) in kinders uit te daag. Akkurate voorspelling oor die uitkoms van TBM is van kritieke belang wanneer doeltreffendheid van verskillende ingrypings beoordeel word. Ek het ‘n retrospektiewe kohort studie van 554 kinders jonger as 13 jaar met TBM wat in Tygerberg Kinderhospitaal toegelaat is oor `n tydperk van twintig jaar (1985 tot 2005) uitgevoer en al die pasiënte volgens die kriteria van al die huidig beskikbare stadiëringsisteme vir kinder TBM geherklassifiseer (hoofstuk 4). Die waarde van die verskillende stadiëringsisteme in die voorspelling van neurologiese uitkoms is toe bepaal. In hierdie studie het ek bevind dat die “Verfynde Mediese Navorsings Raad (MNR) stadiëringsisteem na 1 week” die TBM stadiëringsisteem met die hoogste voorspellende waarde was om neurolgiese uitkoms te voorspel. Dit is geskep deur onderverdeling van stadium 2 (2a en 2b) van die bestaande gemodifiseerde MNR stadiëringsisteem. Daarbenewens het ek ’n vereenvoudigde stadiëringsisteem vir TBM wat minder afhanklik van kliniese vermoëns en neurologiese kundigheid sal wees as die bestaande stadiëringsisteme daargestel en getoets. Die vereenvoudigde stadiëringsisteem is die “Tygerberg Kinderhospitaal Skaal (TKH)” genoem en dit is slegs gebaseer op `n pasiënt se vermoë om visueel te fikseer en te volg en die motoriese respons tot pyn aan beide kante van die ligaam. Dit het uitstekende voorspellingswaarde gehad vir uitkoms na die eerste week van siekte en het in hierdie verband nie betekenisvol verskil van die “Verfynde MNR stadiëringsisteem” nie. Die optimale anti-TB middel regimen en duurte van behandeling vir TBM is onbekend. Sommige kenners stel voor dat ‘n intensiewe kort-kursus (6 maande) van anti-TB behandeling veilig en voldoende mag wees. Ek het ‘n prospektiewe beskrywende studie op 184 opeenvolgende kinders met TBM uitgevoer en bevind dat intensiewe kort-kursus anti-TB behandeling gemik op die behandeling van kinders met TBM (anti-TBM behandeling) in beide menslike immuniteitgebrekvirus (MIV)-ongeïnfekteerde en MIV-geïnfekteerde kinders met middel-gevoelige TBM voldoende en veilig was (hoofstuk 5 ). Die mortaliteit in my studie met voltooing van behandeling vergelyk gunstig met die mediane mortaliteit van 33% (reikwydte 5-65%) wat onlangs in ‘n oorsig van uitkoms in TBM gerapporteer is. TB immuun rekonstitusie inflammatoriese sindrome (IRIS) is ‘n potensieël lewensbedreigende komplikasie in MIV-geïnfekteerde kinders met TB van die sentrale senuwee sisteem (SSS). Min is oor die voorkoms, mortaliteit, onderliggende immunopatologie en behandelingsbenaderings in MIV-geïnfekteerde kinders met neurologiese TB-IRIS bekend. In `n gevalle-reeks het ek gevind dat neurologiese TB-IRIS oorweeg moet word as nuwe neurologiese tekens na aanvang van antiretrovirale terapie (ART) in MIV-geïnfekteerde kinders met TBM ontwikkel (hoostuk 6.1). Simptome en tekens van neurologies TB-IRIS behels hoofpyn, konvulsies, meningiale prikkeling, ‘n verlaagde vlak van bewussyn, ataksie en fokale motoriese uitval. Ons bespreek ook die rasionaal vir die gebruik van sekere behandelingsmodaliteite, insluitende thalidomied. Neurologiese tuberkuleuse massaletsels (tuberkulome en pseudo-absesse) mag ontwikkel of vergroot in kinders op anti-TBM behandeling. Hierdie letsels reageer swak op terapie, vereis soms chirurgiese verwydering, maar kan op talidomied behandeling reageer, ‘n kragtige inhibeerder van tumor nekrose faktor-alfa (TNF-α). Die optimale dosis en duurte van thalidomide behandeling en die korrelasie met magnetiese resonansbeelding (MRB) moet nog ondersoek word. Die primêre doel van my volgende studie was om te bepaal of seriële MRB van waarde is om die respons op behandeling te evalueer asook die duurte van talidomied behandeling. Die sekondêre doelwit was om die waarde van talidomied in die behandeling van hierdie letsels te bepaal. In ‘n prospektiewe waarnemingstudie wat oor 3 jaar gestrek het is seriële MRB uitgevoer op 16 opeenvolgende kinders met TB pseudo-absesse wat behandel is met talidomied (hoofstuk 6.2). Die spoedige kliniese verbetering van die meeste pasiënte dui daarop dat thalidomied `n aansienlike kliniese voordeel bied in hierdie kliniese konteks. Verder het ek `n MRB merker van genesing geïdentifiseer naamlik evolusie van die letsel van vroeë stadium “T2 helder” met edeem na “T2 swart”. Hierdie bevinding is van groot waarde in die toekomstige behandeling van TBM pasiënte wat hierdie komplikasie ontwikkel. Transkraniale Doppler beelding (TKDB) is potensieël `n waardevolle ondersoekmetode in kinders met TBM, `n toestand wat dikwels gekompliseer word deur patologie verwant aan Doppler beelding soos verhoogde intrakraniale druk (IKP) en serebrale vaskulopatieë. Seriële TKBD is op 20 TBM kinders uitgevoer om serebrale hemodinamika en die verband tussen die pulsatiele indeks (PI) en IKP te ondersoek (hoofstuk 6.3). In hierdie studie het ek gevind dat TKDB-afgeleide PI nie `n betroubare aanduiding van verhoogde IKD in kinders met tuberkuleuse hidrokefalus is nie en dit aan individuele variasies van tuberkuleuse vaskulêre siekte toegeskryf, wat serebrale vaskulêre toegeeflikheid en weerstand benadeel. Die studie het die doeltreffendheid van mediese behandeling in kinders met kommunikerende tuberkuleuse hidrokefalus bevestig. In alle gevalle het die IKP binne 7 dae na aanvang van asetosoolamied en furosemied genormaliseer. In dieselfde groep TBM kinders het ek die serebrale bloedvloei-snelhede (BVS) in die anterior serebrale arterie (ASA), middel serebrale arterie (MSA) en posterior serebrale arterie (PSA) met toelating en na dag 3 en 7 gemeet. Ek het volgehoue hoё BVS in al die basale arteries gevind wat op stenose sekondêr tot vaskulitis eerder as funksionele vasospasma dui. Daarbenewens het ek gevind dat volledige MSA afsluiting, subnormale gemiddelde MSA snelhede (minder as 40 sentimeter per sekonde) en `n verminderde PI (minder as 0.4) met radiologies-bewysde groot serebrale infarksies gekorreleer het. Geen kant-tot-kant verskille in MSA BVS of subnormale PI’s is in vier TBM kinders met kleiner infarksies met toelating bespeur nie. Ek skryf dit toe aan die afsluiting van `n beperkte aantal (een of twee) van die nege MSA perforators wat nie nie die hemodinamika van die MSA beïnvloed nie. Ek het afgesluit om al die vrae wat nog bestaan oor die beste benadering ten opsigte van voorkoming, diagnose and behandeling van TBM uit te wys (hoofstuk 2). In die tweede literatuuroorsig, wat gemik is op dokters wat werk in hulpbron-beperkte lande, beskryf ek nuwe benaderings tot die hantering van pediatriese TBM, insluitend `n behandelingsalgoritme vir tuberkuleuse hidrokefalus, die rol van kort- kursus versterkte anti-TB behandeling vir TBM en tuis-gebaseerede anti-TBM behandeling (hoofstuk 3). Selfs met die beste diagnostiese en behandelingsmodaliteite, is die uitkoms van kinder TBM swak indien diagnose vertraag word. Ons pogings moet daarom op groter bewustheid en vroeёr diagnose berus.

Base teórica: A meningite bacteriana é uma causa importante de morbidade e mortalidade na infância. Análise do líquido cefalorraquidiano (LCR) continua a ser a ferramenta de diagnóstico padrão ouro, porém novos biomarcadores para o diagnóstico e prognóstico ainda são necessários. Receptor Desencadeador Expresso nas Células Mielóides Tipo 1 (TREM-1) é um receptor transmembrana expresso em neutrófilos e monócitos, que desempenha um papel importante na modulação da resposta inflamatória. A sua fração solúvel (sTREM-1) também é aumentada na infecção, inflamação ou doenças imunológicas. Neste estudo nós avaliamos, prospectivamente, o valor do TREM-1 como um biomarcador de meningite bacteriana aguda em pacientes pediátricos e sua possível utilização como uma ferramenta de prognóstico neste cenário. Objetivos: O objetivo primário do presente estudo é caracterizar os níveis líquóricos solúveis de TREM-1 (sTREM-1) em pacientes admitidos por suspeita clínica de meningite. Analisamos também os níveis de sTREM-1 nos casos de meningite bacteriana e viral, além de medir a sensibilidade e especificidade deste biomarcador no LCR e estudar se esse biomarcador pode ser um fator associado ao prognóstico em meningite bacteriana aguda. Métodos: Sessenta e um pacientes pediátricos, de 0 a 10 anos foram avaliados quanto à meningite e foram prospectivamente incluídos neste estudo. Na admissão, após a suspeita clínica de meningite foram submetidos à análise do LCR para o diagnóstico e uma amostra do LCR inicial foi utilizado também para análise do sTREM-1. Os pacientes foram acompanhados durante a sua internação com o registro de seu tratamento e desfecho clínico para posterior análise dos dados. Resultados: Dentre os 61 pacientes, 38 (62%) foram negativos para a meningite, 7 (11%) pacientes foram diagnosticados com meningite viral e 16 (27%) pacientes foram diagnosticados com meningite bacteriana aguda e recebeu tratamento direcionado. Sexo (p = 0,15), presença de fatores de risco identificados (p = 0,17), presença de convulsões (p = 0,31), outras complicações clínicas (p = 0,11) e mortalidade (p = 0,66) não diferiram entre os grupos. Anormalidades sensoriais (p <0,0001) e presença de cefaléia (p = 0,003) foram mais prevalentes em pacientes com meningite. Como esperado, a contagem de leucócitos, glicose e proteína no LCR foram significativamente diferentes entre pacientes com meningite e pacientes sem meningite. As concentrações de sTREM-1 no LCR de pacientes com meningite bacteriana foi superior quando comparada com pacientes com meningite viral e com controles (1204,67 pg/ml, 39,34 pg/ml e 12,09 pg/ml, respectivamente; p <0,0001). Quando sTREM-1 foi usado como um determinante de diferenciação entre pacientes com ou sem meningite bacteriana, a análise da área sob a curva ROC foi de 0,95 (IC de 95% = 0,89-1,00; p <0,0001). A presença de fatores de risco para a meningite bacteriana (p = 0,04), anormalidades sensoriais (p <0,0001), contagem de leucócitos no LCR (p = 0,01), níveis de glicose no LCR (p = 0,002), níveis de proteína no LCR (p = 0,032) e os níveis de sTREM-1 no LCR (p = 0,004) foram associados com meningite bacteriana, incluindo os níveis sTREM-1 acima do ponto de corte estabelecido de 68,0 pg/ml (p <0,0001). A meningite bacteriana (p = 0,02) e os valores de sTREM-1 maior do que o ponto de corte (68,0 pg/ml) (p = 0,04) foram associados com sequelas neurológicas graves e morte neste grupo de pacientes. Conclusão: Avaliamos os níveis sTREM-1 de crianças com suspeita clínica de meningite. Os níveis de s-TREM-1 foram aumentados nos casos de meningite bacteriana e correlacionados com o prognóstico. Os nossos resultados sugerem que níveis elevados de sTREM-1 no LCR podem ser utilizados como um biomarcador para o diagnóstico de meningite bacteriana aguda em crianças e que pode ser útil na determinação do prognóstico do paciente nesse cenário.
Background: Bacterial meningitis is an important cause of morbidity and mortality in infancy. Cerebrospinal fluid (CSF) analysis remains the gold standard diagnostic tool, however new biomarkers for diagnosis and prognosis are still required. Triggering receptor expressed on myeloid cells-1 (TREM-1) is a transmembrane receptor expressed on neutrophils and monocytes that plays an important role on the immune response. Its soluble fraction (sTREM-1) is also increased in infection, inflammation or immune diseases. In this study we evaluate the value of sTREM-1 as a biomarker of acute bacterial meningitis in pediatric patients and its possible use as a prognostic tool prospectively. Methods: Sixty-one pediatric patients, from 0 to 10 years of age were evaluated for meningitis and were prospectively included in this study. At admission, following clinical hypothesis of meningitis patients were submitted to CSF analysis for diagnosis and a sample of initial CSF was also used for TREM-1 analysis. Patients were followed during hospitalization and clinical evaluation and treatment outcome were recorded for posterior analysis. Results: Thirty-eight (62%) out of 61 patients were negative for meningitis, 7 (11%) patients were diagnosed with viral meningitis and 16 (27%) patients were diagnosed with and received treatment for acute bacterial meningitis. Sex (p = 0.15), presence of identified risk factors (p = 0.17), presence of seizures (p = 0.31), other clinical complications (p = 0.11), and mortality (p = 0.66) did not differ among groups. Sensorial abnormalities (p<0.0001) and presence of headache (p= 0.003) were more prevalent in patients with meningitis. As expected, leukocyte count, glucose, and protein levels were significantly different between patients with meningitis and patients without meningitis. Concentrations of sTREM-1 in CSF from patients with bacterial meningitis was higher when compared to patients with viral meningitis and with controls (1204.67 pg/ml, 39.34 pg/ml and 12.09 pg/ml, respectively; p<0.0001). When sTREM-1 was used as a determinant to differentiate between patients with or without bacterial meningitis, the analysis of the area under the ROC curve (AUC) was 0.95 (95% CI=0.89-1.00; p<0.0001). Presence of risk factors for bacterial meningitis (p = 0.04), sensorial abnormalities (p<0.0001), CSF leukocyte count (p = 0.01), CSF glucose levels (p = 0.002), CSF protein levels (p = 0.032) and CSF sTREM-1 levels (p = 0.004) were all associated with bacterial meningitis, including sTREM-1 levels above the established cut-off point of 68.0 pg/ml (p<0.0001). Bacterial meningitis (p = 0.02) and values of sTREM-1 higher than the cut-off point (68.0 pg/ml) (p = 0.04) were associated with death and severe neurological disabilities in this patient cohort. Conclusion: We evaluated sTREM-1 levels in CSF of children with clinical hypothesis of meningitis. The sTREM-1 levels were increased in bacterial meningitis and correlated with prognosis. Our results suggest that CSF sTREM- 1 levels can be used as a biomarker for diagnosis of acute bacterial meningitis in children and it might be useful in determining patient’s prognosis in this scenario.

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Fundação Oswaldo Cruz. Centro de Pesquisas Gonçalo Moniz. Salvador, BA, Brasil
INTRODUÇÃO: Em 2010, a vacina conjugada 10-valente (PCV10) foi incorporada ao programa nacional de imunizações (PNI) brasileiro. Este imunobiológico confere imunização contra os dez principais tipos capsulares de Streptococcus pneumoniae, patógeno responsável por diversas manifestações clínicas e com elevada contribuição nas taxas de incidência e mortalidade por meningite, que é a condição clínica mais grave. OBJETIVO: O presente estudo teve como objetivo avaliar o impacto da PCV10 na epidemiologia da meningite pneumocócica na região metropolitana de Salvador (RMS) Bahia, comparando o período anterior (2008-2010) e posterior (2011-2013) a sua utilização, bem como realizar uma caracterização molecular minuciosa a partir de uma série histórica (1996-2012) entre os isolados resistentes a penicilina (PNSSP com CIM≥ 0,125 μg/mL) e para os sorotipos não-vacinais (2008-2012). MATERIAL E MÉTODOS: Foram incluídos todos casos de meningite pneumocócica confirmados laboratorialmente no período entre 1996 a 2013. Taxas de incidência para a Salvador e RMS foram calculadas com base nos dados populacionais do IBGE/2010. A determinação do tipo capsular foi realizada através da técnica de Multiplex-PCR e/ou reação de Quellung. A sensibilidade a nove antimicrobianos foi testada através das técnicas disco-difusão, microdiluição e E-test. Para caracterizar o perfil molecular foram aplicadas as técnicas de genotipagem de PFGE e MLST. RESULTADOS: Um total de 939 casos de meningite pneumocócica foram identificados no período de 1996- 2013, sendo que 70 casos ocorrem entre 2011 a 2013 (período pós-vacinal). A incidência de meningite pneumocócica em todas as faixas etárias na RMS reduziu de 0,70 casos/100.000 habitantes para 0,59 casos/100.000 habitantes considerando o período de três anos antes e após a vacinação com PCV10 [p< 0,05; RR IC 95%: 1,46 (1,03-2,05)]. Esta redução foi significativa na faixa etária de 0-2 anos e nos casos por sorotipos relacionados à PCV10. Não houve aumento significativo de casos por sorotipos não vacinais nesta casuística, apesar do surgimento de casos por sorotipos não-vacinais não detectados anteriormente na série histórica de MP (10F, 21, 22F, 15A e 24F). Os isolados resistentes à penicilina analisados na série histórica se restringiram a 13 sorotipos, entre os quais: 14 (45,1 %; 78/173), 23F (19,1%; 33/173), 6B (14,4 %; 25/173), 19F (9,2 %; 16/173) e 19A (5,2 %; 9/173). 94% dos casos nãosusceptíveis à penicilina (PNSSP) foram de sorotipos vacinais. Os grupos clonais caracterizados pelo PFGE/MLST predominantes ao longo dos anos foram representados pelo sorotipo 14, denominado grupo A/ST 66 [35,3 % (61/173)] e grupo GK/ST 156 [4.6 % (8/173)], este último associado com níveis elevados de resistência a penicilina e ceftriaxona. Não foram detectados grupos clonais emergentes associados a tipos capsulares não-vacinais. CONCLUSÕES: Estes achados sugerem que a introdução da PCV10 modificou a epidemiologia da meningite pneumocócica na população estudada.
INTRODUCTION: In 2010, the 10-valent pneumococcal conjugate vaccine (PCV10) was introduced into the Brazilian national immunization program (NIP). This immunobiological provides immunization against the main ten capsular types of Streptococcus pneumoniae, the pathogen responsible for different clinical manifestations and high contribution in the incidence and mortality from meningitis, which is the most severe clinical condition. OBJECTIVE: This study aimed to evaluate the impact of PCV10 in the epidemiology of pneumococcal meningitis in the metropolitan area of Salvador (RMS) Bahia, comparing the previous (2008-2010) and after (2011-2013) periods its use, as well as conduct a thorough molecular characterization from a historical series (1996-2012) among isolates resistant to penicillin (PNSSP with CIM≥ 0.125 g / ml) and nonvaccine serotypes (2008-2012). MATERIAL AND METHODS: We included all cases of pneumococcal meningitis laboratory confirmed for the period 1996 to 2013. Incidence rates for Salvador and RMS were calculated based on population data from IBGE/2010. The capsular type determination was performed by multiplex PCR and/or Quellung reaction. Isolates Nine antibiotics were tested by disk-diffusion test, broth micro-dilution and E-test. To characterize the molecular profiling techniques were applied genotyping PFGE and MLST. RESULTS: A total of 939 cases of pneumococcal meningitis were identified during 1996-2013 period, with 70 cases occurring between 2011-2013 (post-vaccination period). The incidence of pneumococcal meningitis in all age groups in the RMS decreased from 0.70 cases / 100,000 inhabitants to 0.59 cases / 100,000 inhabitants considering the three-year period before and after vaccination with PCV10 [p <0.05; RR 95% CI: 1.46 (1.03 to 2.05)]. This reduction was significant in the age group 0-2 years and in cases by serotypes related to PCV10. There was no significant increase in cases by serotypes not vaccine in this series, despite the emergence of cases by serotypes not-vaccine previously undetected in the historical series of MP (10F, 21, 22F, 15A and 24F). The penicillin resistant isolates analyzed the historical series were restricted to 13 serotypes, including: 14 (45.1%; 78/173), 23F (19.1%; 33/173), 6B (14.4%; 25/173), 19F (9.2%, 16/173) and 19A (5.2%, 9/173). 94% of nonsusceptible to penicillin cases (PNSSP) were vaccine serotypes. Clonal groups characterized by PFGE / MLST predominant over the years have been represented by serotype 14, group called A / ST 66 [35.3% (61/173)] and Group GK / TS 156 [4.6% (8/173) ], the latter associated with elevated levels of penicillin and ceftriaxone resistance. Not were detected emerging clonal groups associated with capsular types non-vaccination. CONCLUSIONS: These findings suggest that the introduction of PCV10 changed the epidemiology of pneumococcal meningitis in the population studied.

Introdução: O implante auditivo de tronco encefálico (ABI) é indicado para pacientes com perda auditiva neurossensorial severa a profunda quando há impossibilidade de realização do implante coclear. Em pacientes com surdez por meningite e ossificação coclear total bilateral, o ABI é a opção para a reabilitação auditiva. Objetivos: O estudo tem por objetivo avaliar a contribuição do ABI para os limiares audiométricos e para a percepção de fala após no mínimo 12 meses de uso em pacientes com ossificação coclear total por surdez pós-meningite, bem como descrever as complicações do procedimento. Material e métodos: Dez pacientes com surdez pósmeningite foram submetidos ao ABI por via retrolabiríntica ampliada em um centro terciário de assistência e ensino. Todos os pacientes foram operados pela mesma equipe cirúrgica e a avaliação audiológica foi realizada pelo mesmo fonoaudiólogo. Foram realizados audiometria tonal e testes de percepção de fala no pré-operatório e no mínimo 12 meses após a ativação do implante. Foram descritas as complicações associadas ao procedimento. Resultados: Oito de dez pacientes implantados permaneceram usuários. Dois pacientes não apresentaram respostas auditivas e abandonaram o seguimento. Os oito pacientes apresentaram melhora estatisticamente significativa nos limiares audiométricos, bem como nos testes de discriminação de palavras e vogais comparando pré e pós-operatório com média de seguimento de 3,3 anos. Em dois pacientes, a discriminação de sentenças em formato fechado somente no modo auditivo foi de 30% e 40%. Todos os oito usuários referiram benefício com o uso do ABI. Não houve complicações relacionadas ao procedimento. Conclusão: O ABI via retrolabiríntica ampliada é uma opção terapêutica segura para pacientes com surdez pós-meningite e com presença de ossificação coclear total bilateral, contribuindo para melhora nos limiares audiométricos e nos testes de percepção de fala. Embora a melhora nos testes audiológicos seja inferior à do implante coclear, a maioria dos pacientes do estudo usa o ABI diariamente por um período superior a 8 horas e refere benefício em seu cotidiano
Introduction: The Auditory Brainstem Implant (ABI) is an option to auditory restoration in patients with severe to profound sensorineural hearing loss who cannot be fitted with a cochlear implant. This is the only option in patients with post meningitis hearing loss presenting with bilateral total cochlear ossification. Objectives: The main objective of the study is to evaluate the hearing contribution in audiometry and speech perception tests at least 12 months after ABI implantation in patients with post-meningitis profound hearing loss. The complications of the procedure were also described. Materials and Methods: Ten patients with post-meningitis hearing loss went an ABI through extended retrolabyrinthine approach in a tertiary center by the same surgeons. The same audiologist was responsible for audiological follow-up. Tonal audiometry and speech perception tests were made before and at least 12 months after the ABI activation. The procedure complications were described for all patients. Results: Eight of ten patients became ABI users. Two patients had no auditory response and abandoned the treatment. Eight users showed benefit in tonal audiometry, word and vowels perception tests after an average follow up of 3.3 years. Two patients were able to recognize 30 and 40% of closed sentences without lip reading. There were no complications due to the ABI procedure. Conclusion: The extended retrolabyrinthine approach for the ABI is a safe surgical option for patients with post-meningitis hearing loss and totally ossified cochleae. It contributes to hearing performance in audiometry and speech perception tests. Even though the ABI results are poorer than the cochlear implants, in this study the majority of patients use their ABI more than eight hours a day and report benefits in daily activities

Includes bibliographical references.
Acute bacterial meningitis is defined as the inflammation of the meninges. It is caused by various bacteria and the specific aetiology is age dependant. In the neonatal period the causative organisms are: Group B streptococci, Gram - negative bacilli (e.g.: E. coli, Klebsiella spp, Enterobacter spp, Salmonella spp) and Listeria monocytogenes. In infants and children up to the age of 5 the most common causative organisms include: Streptococcus pneumoniae, Haemophilus influenzae type B (Hib)and Neiseria meningitidis. The two chief causes of bacterial meningitis in children older than 5 are S. pneumoniae and N. meningitidis. Various studies have been performed to look at the profile of meningitis among the paediatric population. Objective: To investigate the aetiology of acute bacterial meningitis in South African newborns and children from 2005 - 2010.

Epidemiological data suggest that viral infections might be predisposing factors for bacterial meningitis and carriage of type b Haemophilus influenzae (Hib), Neisseria meningitidis or Streptococcus pneumoniae, the three pathogens most commonly associated with bacterial meningitis. Non-secretors of ABO blood group antigens are over-represented among patients with bacterial meningitis and in populations affected by some outbreaks. The first objective of the study was to examine the hypothesis that non-secretors were also over-represented among patients with respiratory viral infections. Compared with the local population, there was a significantly higher proportion of secretors among patients with disease due to respiratory syncytial virus (RSV), influenza A virus, rhinovirus and ECHO virus. The hypothesis that Le^b and/or H-type 1 antigens present on cells or in body fluids of secretors might be receptor(s) for RSV was examined. Affinity purified molecules with Le^b or H type 1 determinants or synthetic receptor analogues did not decrease the infectivity of RSV for HEp-2 cells. Bacterial attachment to mucosal surfaces is an important prerequisite for infection. The second major objective was to assess the effect of RSV infection of HEp-2 cells (a human epithelial cell line) on binding of bacteria responsible for meningitis. Binding of bacteria labelled with fluorescein isothiocyanate (FITC) to HEp-2 cells and RSV-infected HEp-2 cells was compared by flow cytometry. Strains of meningococci (3) and Hib (5) expressing antigens of different serogroups, serotypes and subtypes and a strain of Staphylococcus aureus bound significantly more effectively to virus infected cells. Similar patterns of increased binding of unlabelled meningococci to monolayers of RSV-infected cells were also observed. Studies to identify the changes on the cell surface associated with RSV infection responsible for enhanced binding were carried out with one strain of meningococcus. Viral infection of HEp-2 cells did not enhance the expression of Lewis^a antigen, a proposed receptor for bacteria in non-secretors. Monoclonal antibodies to the attachment glycoprotein G of RSV decreased the bacterial binding to infected HEp-2 cells; but monoclonal antibodies to the fusion glycoprotein F did not affect binding.